Physiological and Molecular Responses Under Fe Deficiency in Two Rice (Oryza sativa) Genotypes Differing in Iron Accumulation Ability in Seeds

Fe is an essential mineral element that plants need for their growth. When there is low soil availability of Fe, plants show severe deficiency symptoms. Under Fe-deficiency conditions, plants alter a number of processes to acquire Fe from soil. Genes involved in these mechanisms have been identified from different model plants, including Arabidopsis and rice. Fe transport within plants is also tightly regulated. In this study, we used H9405, a cultivar of rice with high Fe accumulation in seeds, and Yangdao 6, a cultivar with low seed Fe accumulation, to study their responses under different Fe conditions. Our results showed that genes involved in acquisition of Fe from soil in these two cultivars were both up-regulated in roots under Fe-deficiency conditions, and the elevation of the expression was much higher in Yangdao 6 than in H9405. However, remobilization-related genes in shoot vasculature were expressed in an opposite way between the two cultivars. In H9405, the expression of these genes was up-regulated; but in Yangdao 6, their expression was reduced. Our results showed that the differential expression of root-uptake and shoot-remobilization genes in the two cultivars is correlated to the Fe content in roots, shoots, and seeds. Strategies to biofortify rice cultivars with different characteristics were also discussed based on our discovery.     

Comparison of cadmium-induced iron-deficiency responses and genuine iron-deficiency responses in Malus xiaojinensis

The effects of the heavy metal Cd in Malus xiaojinensis were investigated using hydroponic cultures. Chlorophyll and Fe concentrations in young leaves were markedly decreased by Cd treatment, although Fe concentration was significantly enhanced in the roots. A comparative examination of the Fe-deficiency responses due to Fe deficiency and Cd treatment was also performed. Both Fe deficiency and Cd treatment induced responses similar to those of Fe-deficiency in M. xiaojinensis, including acidification of the rhizosphere, enhanced Fe(III) chelate reductase activity, and upregulation of the Fe-deficiency-responsive genes MxIRT1 and MxFRO2-Like. However, the Fe-deficiency responses induced by Cd treatment were different in intensity and timing from those induced by Fe deficiency.     

Induced activity of adenine phosphoribosyltransferase (APRT) in iron-deficiency barley roots: a possible role for phytosiderophore production

To isolate the genes involved in the response of graminaceous plants to Fe-deficient stress, a protein induced by Fe-deficiency treatment was isolated from barley (Hordeum vulgare L.) roots. Based on the partial amino acid sequence of this protein, a cDNA (HvAPT1) encoding adenine phosphoribosyltransferase (APRT: EC 2.4.2.7) was cloned from a cDNA library prepared from Fe-deficient barley roots. Southern analysis suggested that there were at least two genes encoding APRT in barley. Fe deficiency increased HvAPT1 expression in barley roots and resupplying Fe to the Fe-deficient plants rapidly negated the increase in HvAPT1 mRNA. Analysis of localization of HvAPT1-sGFP fusion proteins in tobacco BY-2 cells indicated that the protein from HvAPT1 was localized in the cytoplasm of cells. Consistent with the results of Northern analysis, the enzymatic activity of APRT in barley roots was remarkably increased by Fe deficiency. This induction of APRT activity by Fe deficiency was also observed in roots of other graminaceous plants such as rye, maize, and rice. In contrast, the induction was not observed to occur in the roots of a non-graminaceous plant, tobacco. Graminaceous plants generally synthesize the mugineic acid family phytosiderophores (MAs) in roots under Fe-deficient conditions. In this paper, a possible role of HvAPT1 in the biosynthesis of MAs related to adenine salvage in the methionine cycle is discussed.     

缺铁水稻根转录本微点阵分析

水稻不仅是非常重要的粮食作物 ,也是用于研究的模式植物之一 .由于水稻基因组测序的完成 ,用功能基因组学的现代方法来研究缺铁相关基因的表达调控是最高效的方法之一。在前期工作的基础上 ,精心设计了缺铁和EDTA鳌合二价铁诱导5天的水稻根实验 ,并进行了转录水平的微点阵 (microarray)分析。但只获得了第 5天的结果。在 10 5 31个水稻cDNA芯片图谱中 ,缺铁和加铁比较发现了 4 5 1个差异点。对缺铁诱导的 4 5 1个差异cDNA逐一地进行NCBI (美国国家生物技术信息中心 )的BLAST(局部定位排列搜索工具 )数据库检索、分析和归类。发现其中缺铁与加铁 ( -Fe/Fe -EDTAratio)之间的相对表达水平(REL)在 2 - 9.175之间的缺铁诱导上调基因为 2 0 3个 ,缺铁诱导的下调基因为 2 4 8个。对每一类上调基因都逐一地进行了NCBI-PubMed的文献检索。利用国际网络数据库进行了功能鉴定。     

Cadmium inducible Fe deficiency responses observed from macro and molecular views in tobacco plants

Responses induced by Cd exposure were assessed in tobacco seedlings (Nicotiana tabacum L.) using macro and molecular indices. The 100 μM of Cd exposure reduced the total dry weight and chlorophyll index of the seedlings as much as the genuine Fe-deficiency. Concentration of Fe in the shoots decreased, whereas that in the roots increased by the Cd exposure, especially in the apoplasmic space. It is probable that Cd interferes mainly with the step of Fe-translocation from the roots to shoots and this sets the upper-part of the plant in a state of Fe-deficiency. The Cd exposure coordinately increased the expressions of the exogenous and the endogenous Fe-deficiency responsive genes, HvIDS2 _pro ::GUS, NtFRO1 and NtIRT1 in the roots. This is the first data to demonstrate the responses of Cd-inducible Fe-deficiency at a molecular level.     

Nicotianamine synthase gene expression differs in barley and rice under Fe-deficient conditions

Nicotianamine (NA) is an intermediate in the biosynthetic pathway of the mugineic acid family phytosiderophores (MAs), which are crucial components of the iron acquisition apparatus of graminaceous plants. In non-graminaceous plants, NA is thought to be an essential chelator for metal cation homeostasis. Thus NA plays a key role in Fe metabolism and homeostasis in all higher plants. Nicotianamine synthase (NAS, EC 2.5.1.43) catalyzes the trimerization of S-adenosylmethionine to form one molecule of NA. Barley, a plant that is resistant to Fe deficiency, secretes large amounts of MAs, whereas rice, a plant that is susceptible to Fe deficiency, secretes only small amounts. In this study we isolated a genomic fragment containing HvNAS1 from barley and three rice cDNA clones, osnas1, osnas2 and osnas3, from Fe-deficient rice roots. We also isolated a genomic fragment containing both OsNAS1 and OsNAS2. In contrast to barley, in which Fe deficiency induces the expression of NAS genes only in roots, Fe deficiency in rice induced NAS gene expression in both roots and chlorotic leaves. The amounts of endogenous NA in both the roots and leaves were higher than in barley. We introduced barley genomic DNA fragments containing HvNAS1 with either 9 or 2 kb of the 5'-flanking region into rice, using Agrobacterium-mediated transformation. Fe deficiency induced HvNAS1 expression in both roots and leaves of the transgenic rice, as occurs with rice NAS genes. Barley and rice NAS genes are compared in a discussion of alteration of the NAS genes during adaptation to Fe deficiency.     

水稻锌铁转运蛋白ZIP基因家族研究进展

锌(zinc, Zn)和铁(iron, Fe)是水稻(Oryza sativa L.)生长必需的矿质元素,也是人体必需的微量元素。水稻体内Zn、Fe含量维持在适宜水平有利于提高其产量和品质,提高稻米中Zn、Fe含量能够在一定程度上解决人体Zn、Fe营养缺乏的问题。因此,研究水稻中Zn和Fe等微量元素转运蛋白的具体功能对于提高水稻产量和稻米品质具有重要意义。锌铁转运蛋白(zinc-regulated transporters and iron-regulated transporter-like protein, ZIP)负责Zn和Fe等离子的吸收、转运和分配,是维持水稻中Zn和Fe平衡的重要转运蛋白,其表达水平受Zn和Fe水平影响。ZIP基因家族在自然群体中具有丰富的等位变异,而且某些单倍型存在明显的籼粳分化,这可能造成了不同品种间籼、粳稻中Zn和Fe积累的差异。目前,已有大量关于ZIP基因家族的研究,但只有OsZIP3的作用机制研究的较为清楚。另外,对Zn、Fe在籽粒中的积累机制研究和自然群体中ZIP基因的等位变异研究还不够深入。因此,ZIP转运蛋白家族仍存在较大的研究空间。本文详细介绍了ZIP转运蛋白在水稻体内的亚细胞定位、表达模式、转运机制以及在自然群体中的等位变异等,以期为研究水稻稻米微量元素的积累提供理论基础,为提高稻米品质提供借鉴。     

冬小麦植物铁载体分泌的杂种效应

缺铁是石灰性土壤常见的植物营养问题之一.禾本科植物种或基因型的植物铁载体分泌能力与耐缺铁有关,提高植物铁载体分泌能力是改良缺铁的土壤上植物铁aestivum L.) 3个杂交种及其4个亲本在缺铁营养液中植物铁载体的分泌及杂种的效应.植物铁载体的分泌率通过根分泌物对新形成的Fe(OH)3的活化能力进行测定, 在缺铁症出现时每隔2、3天测定1次.在缺铁条件下,所有基因型都分泌较多的植物铁载体,并且随缺铁症状的发展分泌量增加.杂交种具有对缺铁更敏感的反馈系统,在缺铁条件下,杂交种比亲本分泌铁载体的速度更快、量更高.通过分析杂交种和亲本的关系,认为可以通过对亲本分泌植物铁载体能力和配合力的选择,利用杂种优势来提高小麦铁的利用效率.     

Interspecies compatibility of NAS1 gene promoters.

Nicotianamine and nicotianamine synthase (NAS) play key roles in iron nutrition in all higher plants. However, the mechanism underlying the regulation of NAS expression differs among plant species. Sequences homologous to iron deficiency-responsive elements (IDEs), i.e., cis-acting elements, are found on the promoters of these genes. We aimed to verify the interspecies compatibility of the Fe-deficiency response of NAS1 genes and understand the universal mechanisms that regulate their expression patterns in higher plants. Therefore, we introduced the graminaceous (Hordeum vulgare L. and Oryza sativa L.) NAS1 promoter::GUS into dicots (Nicotiana tabacum L. and Arabidopsis thaliana L.). Fe deficiency induced HvNAS1 expression in the shoots and roots when introduced into rice. HvNAS1 promoter::GUS and OsNAS1 promoter::GUS induced strong expression of GUS under Fe-deficient conditions in transformed tobacco. In contrast, these promoters only definitely functioned in Arabidopsis transformants. These results suggest that some Fe nutrition-related trans-factors are not compatible between graminaceous plants and Arabidopsis. HvNAS1 promoter::GUS induced GUS activity only in the roots of transformed tobacco under Fe-deficient conditions. On the other hand, OsNAS1 promoter::GUS induced GUS activity in both the roots and shoots of transformed tobacco under conditions of Fe deficiency. In tobacco transformants, the induction of GUS activity was induced earlier in the shoots than roots. These results suggest that the HvNAS1 and OsNAS1 promoters are compatible with Fe-acquisition-related trans-factors in the roots of tobacco and that the OsNAS1 promoter is also compatible with some shoot-specific Fe deficiency-related trans-factors in tobacco.     

Why are young rice plants highly susceptible to iron deficiency?

The reason why young rice plant is highly susceptible to Fe-deficiency was clarified as follows: Among Gramineae plants rice secreted a very low amount of deoxy-MA as a phytosiderophore even under Fe-deficiency, and the secretion by rice ceased within 10 days under Fe-deficiency although barley secreted MAs during a period of more than one month. When iron depletion continued, the rice root tips become chimeric and epidermal cells became necrotic. The mitochondrial membrane systems in the cortex cells were also severely damaged. Iron starvation occurred even in the mitochondria, and energy charge in the root decreased. This reduced energy charge has firstly diminished the secretion activity of deoxy-MA from the roots, secondly reduced the activity of the transporter which absorb deoxy-MA-Fe^III chelate and finally reduced the synthesis of deoxy-MA from metionine. Consequently, the depletion of Fe^II in the shoot was induced and severe chlorosis rapidly developed in the young rice plant under Fe-deficiency.Abbreviations DCCD dicyclohexylcarbodiimide - CCCP carbonylcyanide-m-chlorophenylhydrazone - MA mugineic acid - MAs mugineic acid-family phytosiderophores, it contains deoxy-MA, MA, epihydroxy-MA, hydroxy-MA, avenic acid and distichonic acid     

In-vitro-cultured subclover root can develop Fe-deficiency stress response

The Fe-deficiency stress response is induced in most plants under Fe-deficient conditions, but whether the shoot and/or the root control development of the stress response is not known. The objectives of the present study were to determine whether in-vitro-cultured subclover roots can develop Fe-deficiency stress response and to examine this approach as a possible screening technique for Fe-deficiency resistance. One-cm long root tips of subclover seedlings were cultured in modified White's medium without (-Fe) or with (+Fe) 100 μM Fe³⁺EDTA. Root Fe³⁺ reduction and H⁺ release were evaluated. On the first day after transfer to the -Fe medium, the Fe-deficiency-resistant cultivar Koala (Trifolium brachycalycinum Katzn. and Morley) started to release H⁺, resulting in a decrease in pH of the culture medium, while the susceptible cultivar Karridale (T. subterraneum L.) did not release H⁺ until the second day. The H⁺-release rate of the -Fe Koala was approximately twice as high as that of the -Fe Karridale for the first 4 days of -Fe treatment. Both Koala and Karridale reached their highest H⁺-release rates on the fourth day after -Fe treatment initiation. The +Fe Koala released H⁺ after several days of culture, but the H⁺ release of the -Fe Koala was severalfold greater than that of the +Fe Koala. The implicit correlation between H⁺ release and Fe-deficiency resistance was substantiated by using a series of subclover cultivars with a range of susceptibilities to Fe deficiency. The pH of the -Fe culture media of the series of cultivars was positively correlated to their Fe-chlorosis scores reported in previous research. The results of the present study indicate that root itself has the full ability to develop Fe-deficiency stress response and the response is dependent on the root Fe status. The results also suggest that root culture could be used as a simple and efficient alternative technique for screening germplasm for Fe-deficiency resistance.     

Expression of iron-acquisition-related genes in iron-deficient rice is co-ordinately induced by partially conserved iron-deficiency-responsive elements

Rice plants (Oryza sativa L.) utilize the iron chelators known as mugineic acid family phytosiderophores (MAs) to acquire iron from the rhizosphere. Synthesis of MAs and uptake of MA-chelated iron are strongly induced under conditions of iron deficiency. Microarray analysis was used to characterize the expression profile of rice in response to iron deficiency at the genomic level. mRNA extracted from iron-deficient or iron-sufficient rice roots or leaves was hybridized to a rice array containing 8987 cDNA clones. An induction ratio of greater than 2.0 in roots was observed for 57 genes, many of which are involved in iron-uptake mechanisms, including every identified or predicted step in the methionine cycle and the biosynthesis of MAs from methionine. Northern analysis confirmed that the expression of genes encoding every step in the methionine cycle is thoroughly induced by iron deficiency in roots, and almost thoroughly induced in leaves. A promoter search revealed that the iron-deficiency-induced genes related to iron uptake possessed sequences homologous to the iron-deficiency-responsive cis-acting elements IDE1 and IDE2 in their promoter regions, at a higher rate than that showing no induction under Fe deficiency. These results suggest that rice genes involved in iron acquisition are co-ordinately regulated by conserved mechanisms in response to iron deficiency, in which IDE-mediated regulation plays a significant role.     

Do radial oxygen loss and external aeration affect iron plaque formation and arsenic accumulation and speciation in rice?

Hydroponic experiments were conducted to investigate the effect of radial oxygen loss (ROL) and external aeration on iron (Fe) plaque formation, and arsenic (As) accumulation and speciation in rice (Oryza sativa L.). The data showed that there were significant correlations between ROL and Fe concentrations in Fe plaque produced on different genotypes of rice. There were also significant differences in the amounts of Fe plaque formed between different genotypes in different positions of roots and under different aeration conditions (aerated, normal, and stagnant treatments). In aerated treatments, rice tended to have a higher Fe plaque formation than in a stagnant solution, with the greatest formation at the root tip decreasing with increasing distances away, in accordance with a trend of spatial ROL. Genotypes with higher rates of ROL induced higher degrees of Fe plaque formation. Plaques sequestered As on rice roots, with arsenate almost double that with arsenite, leading to decreased As accumulation in both roots and shoots. The major As species detected in roots and shoots was arsenite, ranging from 34 to 78% of the total As in the different treatments and genotypes. These results contribute to our understanding of genotypic differences in As uptake by rice and the mechanisms causing rice genotypes with higher ROL to show lower overall As accumulation.